PROJECT SUMMARY/ABSTRACT Multiple sclerosis (MS) is a chronic inflammatory disease characterized by demyelination and degeneration of axons in the central nervous system (CNS). Early in MS and experimental MS (experimental autoimmune encephalomyelitis, EAE) breakdown of the blood-brain barrier (BBB) and angiogenic remodeling initiate and maintain pathogenesis by affording leukocyte infiltration into the CNS. This vascular remodeling, in turn, is strongly influenced by extracellular matrix (ECM) proteins and their endothelial integrin receptors. We hypothesize that one specific integrin, the ???????laminin receptor, plays an important vasculo-protective role during neuroinflammation by promoting BBB integrity and suppressing endothelial inflammation. This hypothesis is supported by several observations. First, laminin is a major component of the vascular basal lamina, and transgenic mice deficient in astrocyte and pericyte laminin show defective BBB integrity. Second, while most blood vessels in the normal brain express only low levels of ???? integrin, endothelial cells in all vessels strongly upregulate this integrin during inflammation. Third, we have recently found that transgenic mice lacking endothelial ?? integrin expression (??-EC-KO mice) show worse clinical disease and increased neuroinflammation in EAE. Fourth, our in vitro studies of brain endothelial cells suggest that ?4 integrin protects both by maintaining vascular integrity (via claudin-5 stabilization) and by reducing vascular activation (via reduced ICAM-1 expression). Taken together, these data suggest that ???? integrin upregulation may be an inducible protective mechanism that stabilizes the BBB under conditions of stress. Furthermore, we have found that the cholesterol-lowering drug atorvastatin upregulates endothelial ?? integrin expression in vivo, suggesting that this could provide an effective way to therapeutically manipulate ?? integrin levels in MS. As part of these studies we will also generate a novel transgenic mouse strain in which ?? integrin is over-expressed specifically in endothelial cells (??-EC-high) and test whether these mice are protected against neuroinflammation. In vitro studies will define which inflammatory mediators induce endothelial ?4 integrin expression and examine the mechanisms underlying statin-induction of endothelial??? integrin. Lastly, we will define the roles of three signaling pathways in mediating ?? integrin-mediated vasculo- protection: (i) TGF-? ALK1-Smad 1/5/8, (ii) MAP kinase, and (iii) myosin light chain (MLC) kinase. Our hypothesis will be tested in three specific aims: (1) define mechanistically how ?4 integrin protects against neuroinflammatory disease, (2) demonstrate that upregulation of endothelial ?4 integrin protects against EAE and (3) define the intracellular?signaling pathways that mediate the vasculo-protective influence of??? integrin. The approach is innovative because it will test the novel concept that ?? integrin promotes BBB integrity and suppresses vascular inflammation during EAE. This research is significant because successful completion of these studies will evaluate endothelial ???? integrin as a novel MS therapeutic target.